The development of new polymeric membranes is an area of intense commercial interest because of their usefulness in many different applications. Membranes can be defined as selective barriers between two phases. Efficient separation is achieved by the differential rate of movement of molecules, and is dependent on the properties of the separation medium, for example, porosity, pore size distribution, thickness, hydrophilicity, membrane fouling, etc. Examples of the driving force for the movement of molecules across membranes includes concentration differences, pressure differences and electric potential difference (e.g., electrophoresis-based systems).
A wide variety of different materials has been utilized for producing membranes. In general, microporous membranes can be divided into two main groups: those formed physically and those formed chemically. Physically formed membranes can be controllably formed by careful manipulation of the solubility of polymers in solution. These physically formed membranes may be produced by diffusion induced phase separation techniques (DIPS) or temperature induced phase separation (TIPS). Physically formed membranes are useful for many applications including water purification, dialysis and protein separation. However, the techniques for reliably producing physically formed membranes of controlled pore size distribution can be complicated, expensive and not easily reproduced in the laboratory.
Chemically produced membranes may be made via a series of chemical reactions to form very thin three-dimensional polymeric networks. Because these thin polymeric networks generally lack mechanical strength, they are often supported by a substrate that provides the membrane with the requisite mechanical strength. Examples of such membranes include those formed from acrylics, vinylics, methyl methacrylates/ethylene glycol dimethacrylate (EGDMA) and acrylamide (AAm)/N,N′-methylene-bis-acrylamide (Bis) networks. Polymer membranes have been formed by free radical chain polymerization. Unfortunately, free radical reactions are difficult to control, resulting in unwanted side reactions and charged groups.
Membranes are utilized in a wide variety of applications, and are particularly useful in electrophoretic techniques. For example, one membrane-based electrophoresis technique (e.g., Gradiflow™ (Gradipore, Australia)) involves a fixed boundary preparative electrophoresis method (U.S. Pat. Nos. 5,650,055, 5,039,386 and WO 0013776). This technique utilizes a semi-permeable membrane to separate two streams of macromolecules—(e.g., proteins, DNA, RNA, etc) containing liquids. When an electric potential is applied across the membrane, charged species tend to move in the direction of one of the electrodes. If the charged species are positively charged, they tend to move towards the negative electrode (cathode), conversely, negatively charged species move towards the positive electrode (anode). Careful selection of the properties of the membrane (e.g., pore size distribution) will facilitate the separation of the desired charged macromolecules. Cooling of the solutions is accomplished by circulation of chilled buffer solutions that are separated by two further membranes, hereafter referred to as restriction membranes, and are situated between the electrodes and the separation membranes. The restriction membranes allow the passage of ions but not macromolecules.
Depending on the choice of separation apparatus, separation media, and buffer characteristics, electrophoretic techniques can be used in one or more of at least four different modes: (1) charged-based separation, (2) size-based separation, (3) concentration, and (4) dialysis. There are electrophoresis separation techniques available that can separate compounds on the basis of only one mode whereas the Gradiflow™ is adaptable for separation in each of all four modes by selecting appropriate separation media and electrophoresis conditions.
Hydrogel membranes are currently used in some existing electrophoretic systems. For example, the Gradiflow™ method utilizes a thin polyacrylamide (PAAm) hydrogel membrane with a defined pore size (D. B. Rylatt, M. Napoli, D. Ogle, A. Gilbert, S. Lim, and C. H. Nair, J. Chromatog., A, 865, 145–153, 1999). The membrane is produced via the free radical co-polymerization of a monomer such as acrylamide (AAm) and a polyfunctional crosslinking agent such as N,N′-methylene-bis-acrylamide (Bis). In general, hydrogels are desirable because they are reasonably strong, flexible, chemically inert, bio-compatible and can be made with relatively controlled pore structure for most applications.
Recent work has facilitated advances into producing other polymeric networks as well as improved PAAm gels. One approach has focused on altering the nature of the monomers used, including changing the polyfunctional crosslinking agent, for example in the case of PAAm gels, substitution of Bis for another monomer can lead to a different network structure.(M. G. Harrington and T. E. Zewert, Electrophoresis, 15, 195–199, 1994; G. Y. N. Chan, P. A. Kambouris, M. G. Looney, G. G. Qiao, and D. H. Solomon, Polymer, 41, 27–34, 2000; G. Patras, G. G. Qiao, and D. Solomon, H., Electrophoresis, 21, 3843–3850, 2000). However, due to the free radical nature of the polymerization, the chemistries involved are difficult to control and often result in undesirable defects in the gel. For example, failure to control the reaction conditions of the polymerization can lead to charged groups within the network and reduced stability, thereby decreasing the yield of the reaction and increasing the costs of producing a suitable gel.
Currently, the pore size range of commercially available membranes is somewhat limited. For example, large pores suitable for DNA and RNA separations are not routinely available. Some of the unsolved problems remaining with conventional electrophoresis membranes include producing membranes with no or an insignificant degree of charged groups, the ability to control pore size over a wide range of pore sizes and the development of stable gels over a wide pH range.
Thus, a need exists for polymeric membranes with increased stability, decreased number of charge groups within the gel, that are cost efficient to make, and can be manufactured with increased production yields. It would therefore be beneficial to develop polymeric membranes having one or more properties such as controllable pore sizes, good processability, reproducability, high resistance to degradation, bio-stability and bio-compatibility, and preferably, without one or more disadvantages of existing systems.